The formation of proper synaptic connections among the one hundred billion neurons of the human brain is an extraordinary developmental feat, yet the mechanisms that underlie it are poorly understood. Given the neural complexity of many vertebrates, a simpler vertebrate nervous system is necessary. The proposed project will investigate the mechanisms underlying the development of afferent neuron connectivity in the zebrafish posterior lateral line (PLL). First, the activity-dependence of afferent neurons to form synapses with hair cells of a single polarity will be tested by inhibiting action potentials with tetrodotoxin. Second, by comparing the transcriptomes of anteriorly and posteriorly polarized hair cells, the hypothesis will be tested that hair cells of different polarities have distinct chemical identities that mediate the synapse targeting of afferent neurons. Finally, transgenic zebrafish expressing the Brainbow construct in PLL afferent neurons will be imaged and the lateral line nerve traced. The resulting data will reveal the amount of variability in afferent wiring of the PLL; the process by which PLL afferent neurons are endowed with polarity selectivity; and the organization of afferent synapses at the hindbrain. The results of the proposed research may advance treatments for neurodevelopmental diseases, sensory deficits, and other nervous system disorders.